GLUCONEOGENESIS, GLYCOGEN SYNTHESIS

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GLUCONEOGENESIS, GLYCOGEN SYNTHESIS DEGRADATION

• MIA KUSMIATI
• Departemen BIOKIMIA FK UNISBA

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Overview of gluconeogenesis

• The stimulation of gluconeogenesis by high energy
  charge and high concentrations of citrate and
  acetyl-CoA is counterintuitive.
• Gluconeogenesis is active in the fasting state.
• the energy for gluconeogenesis is supplied by
  fatty acid oxidation.
• During overnight fast 90 gluconeogenesis
  hepar, 10 gluconeogenesis kidney
• Prolonged fasting kidney becomes major glucose
  producing organ (40 total glucose production)

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Overview

• Synthesis of glucose from pyruvate utilizes many
  of the same enzymes as Glycolysis.
• Three Glycolysis reactions have such a large
  negative DG that they are essentially
  irreversible.
• Hexokinase (or Glucokinase)
• Phosphofructokinase
• Pyruvate Kinase.
• These steps must be bypassed in Gluconeogenesis.
• Two of the bypass reactions involve simple
  hydrolysis reactions.

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The reciprocal regulation of glycolysis and
gluconeogenesis in the liver.

• (1), Glucokinase
• (2), phosphofructokinase
• (3), pyruvate kinase
• (4), pyruvate carboxylase
• (5), phosphoenolpyruvate
• (PEP)-carboxykinase
• (6), fructose-1,6-bisphosphatase
• (7), glucose-6-phosphatase
• STIMULATION

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• INHIBITION
• A, Substrate flow during fasting and in the
  well-fed state, and the effects of hormones on
  the amounts of glycolytic and gluconeogenic
  enzymes.
• Regulation of enzyme synthesis and degradation
  is the most important long-term (hours to days)
  control mechanism. In most cases, the hormone
  acts by changing the rate of transcription
  (insulin)

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• B, Short-term regulation of glycolysis and
  gluconeogenesis by reversibly binding effectors
  and by
• - Phosphorylation/dephosphorylation
• - Allosteric and competitive effects
• - phosphorylation.
• Only pyruvate kinase and phosphofructo-2-kinase
  /fructose-2,6-bisphosphatase are regulated by
  cAMP-dependent phosphorylation.

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Regulation of glycolisis gluconeogenesis

• Synthesis and degradation of fructose-2,6-bisphosp
  hate, the most important regulator of
  phosphofructokinase and fructose-1,6-bisphosphatas
  e.
• This regulatory metabolite is synthesized and
  degraded by a bifunctional enzyme that combines
  the kinase and phosphatase activities on the same
  polypeptide.

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Regulation of glycolisis gluconeogenesis

• cAMP-induced phosphorylation inhibits the kinase
  activity and stimulates the phosphatase activity
  of the bifunctional enzyme. , Phosphorylation
  , dephosphorylation , allosteric effect ,
  stimulation , inhibition

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SUBSTRAT for gluconeogenesis

• Lactat
• Pyruvate
• Glycerol
• ?lfa keto acid (oxaloacetat, a ketoglutarat)

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RX Unique to gluconeogenesis

• 7 glycolytic Rx are irreversible are used
  in the synthesis of glucose from lactat or
  pyruvate
• Carboxylation of pyruvate
• biotin is a coenzyme
• Allosteric regualtion
• B. Transport of oxaloacetate to the cytosol
• C. Decaboxylation of cytosolic oxaloacetate
• D. Dephosporilation of Fructose 1,6 biP ?
  fructose 6P
• E. Isomerisasi Fructose 6P? Glucose 6P
• F. Convert glucose 6P ? free glucose

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• Hexokinase or Glucokinase (Glycolysis) catalyzes
• glucose ATP ? glucose-6-phosphate ADP
• Glucose-6-Phosphatase (Gluconeogenesis)
  catalyzes
• glucose-6-phosphate H2O ? glucose Pi

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• Glucose-6-phosphatase enzyme is embedded in the
  endoplasmic reticulum (ER) membrane in liver
  cells.
• The catalytic site is found to be exposed to the
  ER lumen. Another subunit may function as a
  translocase, providing access of substrate to the
  active site.

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• Phosphofructokinase (Glycolysis) catalyzes
• fructose-6-P ATP ? fructose-1,6-bisP ADP
• Fructose-1,6-bisphosphatase (Gluconeogenesis)
  catalyzes
• fructose-1,6-bisP H2O ? fructose-6-P Pi

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• Bypass of Pyruvate Kinase
• Pyruvate Kinase (last step of Glycolysis)
  catalyzes
• phosphoenolpyruvate ADP ? pyruvate ATP
• For bypass of the Pyruvate Kinase reaction,
  cleavage of 2 P bonds is required.
• DG for cleavage of one P bond of ATP is
  insufficient to drive synthesis of
  phosphoenolpyruvate (PEP).
• PEP has a higher negative DG of phosphate
  hydrolysis than ATP.

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• Bypass of Pyruvate Kinase (2 enzymes)
• Pyruvate Carboxylase (Gluconeogenesis) catalyzes
• pyruvate HCO3- ATP ? oxaloacetate ADP
  Pi
• PEP Carboxykinase (Gluconeogenesis) catalyzes
• oxaloacetate GTP ? PEP GDP CO2

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• Contributing to spontaneity of the 2-step
  process
• Free energy of one P bond of ATP is conserved in
  the carboxylation reaction.
• Spontaneous decarboxylation contributes to
  spontaneity of the 2nd reaction.
• Cleavage of a second P bond of GTP also
  contributes to driving synthesis of PEP.

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Pyruvate Carboxylase uses biotin as prosthetic
group.

• Biotin has a 5-C side chain whose terminal
  carboxyl is in amide linkage to the e-amino group
  of an enzyme lysine.
• The biotin lysine side chains form a long
  swinging arm that allows the biotin ring to swing
  back forth between 2 active sites.

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• Biotin carboxylation is catalyzed at one active
  site of Pyruvate Carboxylase.
• ATP reacts with HCO3- to yield carboxyphosphate.
• The carboxyl is transferred from this P
  intermediate to N of a ureido group of the
  biotin ring. Overall
• biotin ATP HCO3- ? carboxybiotin ADP Pi

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• At the other active site of Pyruvate Carboxylase
  the activated CO2 is transferred from biotin to
  pyruvate
• carboxybiotin pyruvate
• ?
• biotin oxaloacetate

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Pyruvate Carboxylase (pyruvate ? oxaloactate)
is allosterically activated by acetyl CoA.
Oxaloacetate tends to be limiting for Krebs
cycle.

• When gluconeogenesis is active in liver,
  oxaloacetate is diverted to form glucose.
  Oxaloacetate depletion hinders acetyl CoA entry
  into Krebs Cycle. The increase in acetyl CoA
  activates Pyruvate Carboxylase to make
  oxaloacetate.

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Carbohydrate Is Stored as Glycogen

• The main stores of glycogen in the body
• Liver? to mantain the blood glucose level
• Skeletal muscle?to serve as a fuel reserve for
  synthesis of ATP during muscle contraction

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STRucture of glycogen

• Glycogen is a branched polymer of between 10,000
  and 40,000 glucose residues held together by
  a-1,4 glycosidic bonds

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STRucture of glycogen
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Synthesis of uridine diphosphate (UDP)-glucose.
UDP-glucose is the activated form of glucose for
glycogen synthesis, but also for the synthesis of
other complex carbohydrates
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• Glucose-6-phosphate is isomerized to
  glucose-1-phosphate by phosphoglucomutase.
• Glucose-1-phosphate then reacts with uridine
  triphosphate (UTP) to form UDP-glucose.
• UDP is attached to C-1 of glucose, and it is
  therefore this carbon that forms the glycosidic
  bond. The bond between glucose and UDP is energy
  rich

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The glycogen phosphorylation reaction.
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• Metabolic fates of glycogen in the liver (A) and
  in muscle (B). Note that the liver possesses
  glucose-6-phosphatase, which forms free glucose
  both in gluconeogenesis and from glycogen. This
  enzyme is not present in muscle tissue.

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• Glycogen breakdown serves different purposes in
  liver and muscle.
• The liver synthesizes glycogen after a
  carbohydrate meal and degrades it to free glucose
  during fasting.
• The glucose-6-phosphate from glycogen breakdown
  is cleaved to free glucose by glucose-6-phosphatas
  e.
• The liver releases this glucose into the blood
  for use by needy tissues, including brain and
  blood cells

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• Skeletal muscle synthesizes glycogen at rest and
  degrades it during exercise.
• Muscles cannot produce free glucose because they
  have no glucose-6-phosphatase.
• Because glycogen degradation produces
  glucose-6-phosphate without consuming any ATP,
  anaerobic glycolysis from glycogen produces three
  rather than two molecules of ATP for each glucose
  residue.

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Glycogen Metabolism Is Regulated by Hormones and
Metabolites

• The phosphorylation state of the enzymes is
  regulated by hormones and their second
  messengers.
• Insulin stimulates glycogen synthesis both in the
  liver and in skeletal muscle. It ensures that
  excess carbohydrate is stored away as glycogen
  after a meal.
• Glucagon stimulates glycogen degradation in liver
  but not muscle during fasting when the blood
  glucose level is low.
• Norepinephrine and epinephrine are powerful
  activators of glycogen breakdown both in muscle
  and liver. They mobilize glycogen when glucose is
  needed to fuel muscle contraction.

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KETERANGAN

• A, Hormonal effects on the phosphorylation of the
  glycogen-metabolizing enzymes by protein kinases
  in the liver. ER, endoplasmic reticulum GSK3,
  glycogen synthase kinase-3
• B, Hormonal effects on the dephosphorylation of
  the glycogen-metabolizing enzymes by protein
  phosphatase-1, and the effects of allosteric
  effectors.

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REGUlation of glycogen metabolism in hepar

• Note that the hormones affect glycogen synthase
  and glycogen phosphorylase through the protein
  kinases and the protein phosphatase
  (phosphatase-1) that regulate their
  phosphorylation state. , Allosteric effects ,
  phosphorylation , dephosphorylation ,
  activation , inhibition.